P601: TP53 ANALYSIS AND REPORTING IN CHRONIC LYMPHOCYTIC LEUKAEMIA: ARE LABORATORIES IN COMPLIANCE WITH THE EUROPEAN RESEARCH INITIATIVE ON CLL (ERIC) RECOMMENDATIONS?
Authors: A. Cartwright, S. Scott, L. Whitby
Journal: HemaSphere (2022)

Background: The presence of TP53 aberrations, specifically del(17p) and/or TP53 nucleotide variants, in CLL are associated with adverse clinical outcomes and resistance to immunochemotherapy. As such, analysis of TP53 has become part of routine clinical diagnostics prior to initial and subsequent treatments, ensuring appropriate risk stratification and therapeutic intervention. Deletions of TP53 are identified by fluorescent in situ hybridisation, however, TP53 variants can be identified through various molecular techniques, including Sanger sequencing and next-generation sequencing (NGS). ERIC recommendations outline the requirements for testing, analysis and reporting of TP53 variants in CLL. Aims: The aim of this study was to evaluate laboratory approaches to analysis and reporting of TP53 variants in CLL in line with ERIC recommendations. Methods: The UK National External Quality Assessment Service for Leucocyte Immunophenotyping has provided external quality assessment for use of NGS gene panels in CLL since 2018. Samples are manufactured from cell line material and one sample per annum is issued for participating laboratories to evaluate using their in-house methodology. Between 2018-2021, three samples were formulated to contain TP53 variants and distributed. Laboratories were requested to report methodology and variants of clinical significance, in line with HGVS nomenclature at both the DNA and protein level. Additionally, details of assay limit of detection (LOD), reference sequences and interpretation databases utilised were also requested. Data returns were then evaluated relating to TP53 variant analysis and reporting. Results: In total, 34 (100%) participating laboratories reported the use of either amplicon or capture based NGS methods, with assay LOD ranging from 1-15%, with a median LOD of 5%. All participating laboratories sequenced the ‘minimum’ exons indicated in the guidelines (exons 4-10), with 91.2% (31) laboratories sequencing ‘optimal’ exons 2-11. Furthermore, 97.1% (33) laboratories utilised the recommended Locus Reference Genomic sequence LRG_321t1 (NM_000546), with one laboratory reporting use of LRG_321t3 (NM_001126114) variant transcript. The IARC TP53 locus-specific database for variant interpretation was utilised by 70.6% (24) laboratories and despite recommendations, use of dbSNP for filtering of polymorphic/neutral variants was undertaken by 76.5% (26) laboratories. Across the 3-sample period, 97.1% (33) laboratories reported correct DNA HGVS nomenclature and 58.8% (20) laboratories reported correct protein HGVS nomenclature. Of the 14 laboratories reporting inaccurate protein HGVS nomenclature, 11 (78.6%) reported minor nomenclature issues precluding full compliance. Summary/Conclusion: In this study, there is a high level of consensus amongst laboratories in the reporting and analysis of TP53 variants, except for protein nomenclature usage. Guidelines serve to promote harmonisation and standardisation of processes within laboratories and ERIC recommendations have achieved this in relation to TP53 variants in CLL, ensuring that patients are appropriately stratified for correct therapeutic interventions. Whilst there is currently a high level of consensus seen in analysis and reporting, the recommendations are likely to evolve as evidence and understanding of the genetic landscape in CLL therapeutics increases and further technological advances are made. As such, laboratories will need to ensure continued adoption to reflect new data reporting methods, treatments and standards of care for CLL patients.